Autoinflammatory mutation in NLRC4 reveals an LRR-LRR oligomerization interface

Publication date: Available online 17 May 2018
Source:Journal of Allergy and Clinical Immunology
Author(s): Fiona Moghaddas, Ping Zeng, Yuxia Zhang, Heike Schützle, Sebastian Brenner, Sigrun R. Hofmann, Reinhard Berner, Yuanbo Zhao, Bingtai Lu, Xiaoyun Chen, Li Zhang, Suyun Cheng, Stefan Winkler, Kai Lehmberg, Scott W. Canna, Peter E. Czabotar, Ian P. Wicks, Dominic De Nardo, Christian M. Hedrich, Huasong Zeng, Seth L. Masters
BackgroundMonogenic autoinflammatory disorders are characterised by dysregulation of the innate immune system, for example by gain-of-function mutations in inflammasome forming proteins such as NLRC4.ObjectiveHere we investigate the mechanism by which a novel mutation in the leucine rich repeat (LRR) domain of NLRC4 (c.G1965C, p.W655C) contributes to autoinflammatory disease.MethodsWe studied two unrelated patients with early onset macrophage activation syndrome (MAS) harboring the same de novo mutation in NLRC4. In vitro, inflammasome complex formation was quantified using flow cytometric analysis of ASC specks. CRISPR/Cas9 techniques and lentiviral transduction were used to generate THP-1 cells with either wild-type or mutant NLRC4 cDNA. Cell death and release of IL-1β/IL-18 were quantified by flow cytometry and ELISA respectively.ResultsThe c.G1965C/p.W655C NLRC4 mutation caused increased ASC speck formation, caspase-1-dependent cell death, and IL-1β/IL-18 production. ASC contributed to p.W655C NLRC4 mediated cytokine release, but not cell death. Mutation of p.W655 activated the NLRC4 inflammasome complex by engaging with two interfaces on the opposing LRR domain of the oligomer. One key set of residues (p.D1010, p.D1011, p.L1012 and p.I1015) participated in LRR-LRR oligomerization when triggered by mutant NLRC4 or T3SS effector (PrgI) stimulation of the NLRC4 inflammasome complex.ConclusionThis is the first report of a mutation in the LRR domain of NLRC4 causing autoinflammatory disease. c.G1965C/p.W655C NLRC4 increased inflammasome activation in vitro. Data generated from various NLRC4 mutations provides evidence that the LRR-LRR interface has an important, previously unrecognized role in oligomerization of the NLRC4 inflammasome complex.

Graphical abstract

Teaser

Two patients with macrophage activation syndrome and elevated serum IL-18 levels were found to have a novel gain-of-function mutation in the leucine rich repeat domain of NLRC4.


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